Analysis of the role of detergent mixtures on the crystallization of the reaction center of Photosystem II (original) (raw)
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The influence of poly(ethylene glycol) (PEG) polymers H-(O-CH2-CH2)p-OH with different average molecular sizes p on the micelle formation of n-alkyl-β-D-maltoside detergents with the number of carbon atoms in the alkyl chain ranging from 10 to 12 is investigated with the aim to learn more about the detergent behavior under conditions suitable for the crystallization of the photosynthetic pigment-protein complex photosystem II. PEG is shown to increase the critical micelle concentration (CMC) of all three detergents in the crystallization buffer in a way that the free energy of micelle formation increases linearly with the concentration of oxyethylene units (O-CH2-CH2) irrespective of the actual molecular weight of the polymer. The CMC shift is modeled by assuming for simplicity that it is dominated by the interaction between PEG and detergent monomers and is interpreted in terms of an increase of the transfer free energy of a methylene group of the alkyl chain by 0.2 kJ mol–1 per 1 ...
Micelle formation in the presence of photosystem I
Biochimica et Biophysica Acta (BBA) - Biomembranes, 2008
The correlation between membrane protein solubilisation and detergent aggregation in aqueous solution is studied for a series of n-alkyl-β-D-maltosides (C x G 2 with x = 10, 11, 12 being the number of carbon atoms in the alkyl chain) using the trimeric photosystem I core complex (PSIcc) of oxygenic photosynthesis from Thermosynechococcus elongatus as model protein. While protein solubilisation is monitored via the turbidity of the solution, the aggregation behavior of the detergent is probed via the fluorescence spectrum of the polycyclic aromatic hydrocarbon pyrene. In addition, changes of the fluorescence spectrum of PSIcc in response to formation of the detergent belt surrounding its hydrophobic surface are investigated. Solubilisation of PSIcc and aggregation of detergent into micelles or belts are found to be strictly correlated. Both processes are complete at the critical solubilisation concentration (CSC) of the detergent, at which the belts are formed. The CSC depends on the concentration of the membrane protein, [prot], and is related to the critical micelle concentration (CMC) by the empirical law ln(CSC/CMC) = n0 [prot], where the constant n0 = (2.0 ± 0.3) μM −1 is independent of the alkyl chain length x. Formation of protein-free micelles below the CSC is not observed even for x = 10, where a significant excess of detergent is present at the CSC. This finding indicates an influence of PSIcc on micelle formation that is independent of the binding of detergent to the hydrophobic protein surface. The role of the CSC in the optimisation of membrane protein crystallisation is discussed. j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / b b a m e m
Focus on the aggregation processes of Photosystem II complexes
Bioelectrochemistry, 2007
In this work the effect of temperature and n-dodecyl-β-D-maltoside (DM) on PSII complexes organization was investigated. An aggregation process of PSII monomers and dimers was documented at different temperatures and low DM concentration by steady-state fluorescence, absorption, circular dichroism, Rayleigh and dynamic light-scattering experiments. Measures of oxygen evolution enabled us to estimate the change in photoactivity of PSII during the aggregation. This process was found to be extensively reversed by increasing DM concentration as proved by means of steady-state fluorescence and dynamic light-scattering experiments.
Spectral, Photophysical, and Stability Properties of Isolated Photosystem II Reaction Center
PLANT PHYSIOLOGY, 1988
Photosystem II reaction center (RC) preparations isolated from spinach (Spinacea oleracea) by the Nanba-Satoh procedure (O Nanba, K Satoh 1987 Proc Nati Acad Sci USA 84: 109-112) are quite labile, even at 4°C in the dark. Simple spectroscopic criteria were developed to characterize the native state of the material. Degradation of the RC results in (a) blueshifting of the red-most absorption maximum, (b) a shift of the 77 K fluorescence maximum from-682 nm to-670 nm, and (c) a shift of fluorescence lifetime components from 1.3-4 nanoseconds and >25 nanoseconds to-6-7 nanoseconds. Fluorescence properties at 77 K seem to be a more sensitive spectral indicator of the integrity of the material. The >25 nanosecond lifetime component is assigned to P680+ Pheophytinrecombination luminescence, which suggests a correlation between the observed spectral shifts and the photochemical competence of the preparation. Substitution of lauryl maltoside for Triton X-100 immediately after RC isolation stabilizes the RCs and suggests that Triton may be responsible for the instability.
Crystallization of the photosystem I reaction centre
The EMBO Journal, 1987
Communicated by J.Jansonius-60 light-harvesting pigment molecules, a number which can vary between different PSI preparations. A third electron acceptor termed X, a (4Fe-4S) iron-sulphur centre, may also be bound by the 65-kd polypeptide (Golbeck and Cornelius, 1986; Hoj and Moller, 1986). There are strong reasons to suspect that the 65-kd band seen on SDS-PAGE is actually composed of two different polypeptides of similar mol. wt and with high amino acid sequence homology (see Fish et al., 1985 for a discussion of this problem). It is likely that the two lower mol. wt polypeptides are involved in the binding of the (4Fe-4S) iron-sulphur acceptors called A and B (Takahashi et al., 1982; R.C.Ford, P.Setif and K.Brettel, unpublished results). Apart from binding electron transfer and light-harvesting components, the PSI reaction centre must interact with other proteins such as the water-soluble electron transfer proteins ferredoxin and plastocyanin as well as membrane-bound, lightharvesting proteins. Clearly, the PSI reaction centre must carry out a variety of functions, but perhaps the most intriguing pro
Photosynthesis Research, 1991
Low temperature (4.2 K) absorption and hole burned spectra are reported for a stabilized preparation (no excess detergent) of the photosystem II reaction center complex. The complex was studied in glasses to which detergent had and had not been added. Triton X-100 (but not dodecyl maltoside) detergent was found to significantly affect the absorption and persistent hole spectra and to disrupt energy transfer from the accessory chlorophyll a to the active pheophytin a. However, Triton X-100 does not significantly affect the transient hole spectrum and lifetime (1.9 ps at 4.2 K) of the primary donor state, P680". Data are presented which indicate that the disruptive effects of Triton X-100 are not due to extraction of pigments from the reaction center, leaving structural perturbations as the most plausible explanation. In the absence of detergent the high resolution persistent hole spectra yield an energy transfer decay time for the accessory Chl a Qy-state at 1.6 K of 12 ps, which is about three orders of magnitude longer than the corresponding time for the bacterial RC. In the presence of Triton X-100 the Chl a Qv-state decay time is increased by at least a factor of 50.
Characterization of a Photosystem II core and its three-dimensional crystals
Photosynthesis Research, 1993
A photosystem II core from spinach containing the chlorophyll-binding proteins 47 kDa, 43 kDa, the reaction center proteins D 1, D2 and cytochrome b m and three low molecular weight polypeptides (MW < 10 kDa) was isolated, its three-dimensional crystals were prepared, and both core and crystals were studied by spectroscopic techniques and electron microscopy. The absorption spectra of the crystallized form of the core indicate a specific orientation of the various pigments within the crystal.
Detergent–protein interactions in aqueous buffer suspensions of Photosystem I (PS I)
Journal of Colloid and Interface Science, 2011
Systematic and uniform monolayer formation of Photosystem I (PS I) onto self-assembled monolayer (SAM) substrates to enable unidirectional electron transfer is crucial for its successful use in the fabrication of bio-hybrid solid-state electronic or photovoltaic devices. Yet, our recent studies indicate that surface self-assembly of PS I from aqueous buffer suspensions onto alkanethiolate SAM/Au substrates frequently leads to complex columnar structures due to solution phase protein aggregations. We investigate the effect of two prototypical non-ionic detergents, n-Dodecyl-b-D-Maltoside (DM) and Triton X-100 (TX-100), on protein-protein interactions via the protein-detergent interfacial chemistry. Dynamic light scattering (DLS) experiments are used to demonstrate the impact of relative protein/detergent concentrations on aggregation dynamics of PS I suspensions. In turn, the surface attachment characteristics of PS I adsorbed from the aforementioned suspensions onto SAM/Au substrate is examined by atomic force (AFM) microscopy. Our results indicate that relative concentration of PS I and detergents (DM or, TX-100) with respect to their critical micelle concentrations (CMC) determines the extent of self-association between PS I complexes driven by the screening induced by detergent micelles and/or, inter-protein distances. Such interfacial phenomena during the PS I-detergent complexation process drives the colloidal system through various regimes of phase separations, suspension and/or, deaggregation, wherein individual PS I complexes can exist in a frustrated state that prevents favorable orientations for PS I-PS I interactions. The present study presents a novel strategy, heretofore not considered, for tailoring inter-protein distances and protein-protein interactions in solution phase, thereby allowing a superior control on the surface attachment of PS I onto SAM/Au substrates.
PLANT PHYSIOLOGY, 1989
The photosystem 11 reaction center as isolated (O Nanba, K Satoh [1987] Proc Nati Acad Sci USA 84: 109-112) is quite dilute and very unstable. Precipitating the complex with polyethylene glycol and resuspending it in buffer without detergent concentrates the reaction center and greatly improves its stability at 40C in the dark as judged by light-induced electron transport activity. Furthermore, a procedure was developed to minimize photodestruction of polyethylene-glycol-concentrated material at room temperature in the light. The ability to stabilize the photosystem 11 reaction center should facilitate future photophysical, biochemical, and structural studies of the complex. The similarity between the primary structures of the L-M dimer of the bacterial reaction center and the D I-D2 proteins of PSII (19, 25) suggested an analogous structural and functional relationship between the reaction center of both types of organisms (7, 22). This suggestion was recently supported by the isolation of the Dl-D2-Cyt b-559 complex from spinach (17) and, more recently, pea (1). This complex contains four to five chlorophylls, two Pheos,5 one A-carotene, and one to two heme prosthetic groups per reaction center, but it lacks quinones and Mn (17). Nevertheless, photophysical studies demonstrated that the complex was the simplest PSII preparation yet isolated that still performs the following primary photochemical step (6): h,v